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Hi-C ku salaysan isdhexgalka Chromatin
Hi-C waa hab loogu talagalay in lagu qabto qaabeynta genomic iyadoo la isku darayo isdhexgalka ku saleysan dhowaanshaha iyo isku xigxiga-sare. Habka waxa uu ku salaysan yahay iskutallaabta chromatin ee leh formaldehyde, oo ay ku xigto dheefshiidka iyo dib-u-celinta qaab kaliya jajabyada si wadajir ah ugu xiran ayaa samayn doona alaabooyin ligation ah. Iyadoo la raacayo alaabooyinkan ligation, waxaa suurtagal ah in la barto ururka 3D ee genome. Hi-C waxa ay sahlaysa in la barto qaybinta qaybaha genome-ka kuwaas oo si fudud loo soo buuxiyey (Qaybyada A, euchromatin) oo ay u badan tahay in ay si qoraal ah u firfircoon yihiin, iyo gobollada si adag u xidhxidhan (Qaybaha B, Heterochromatin). Hi-C waxa kale oo loo isticmaali karaa in lagu tilmaamo Domains Topologically Associated (TADs), gobollada genome ee leh qaabab isku laabma oo ay u badan tahay in ay leeyihiin qaabab hadal isku mid ah, iyo in la aqoonsado wareegyada chromatin, gobollada DNA ee ay isku dhejiyeen borotiinno iyo kuwan inta badan lagu hodmay xubno nidaamsan. Adeegga isku xigxiga ee BMKGene ee Hi-C waxa uu awood u siiyaa cilmi-baarayaasha in ay sahamiyaan cabbirrada baaxadda genomics-ka, iyaga oo furaya waddooyin cusub oo lagu fahmo nidaamka genome-ga iyo saamaynta ay ku leedahay caafimaadka iyo cudurrada.
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Chromatin Taxanaha Tallaalka Tallaalka (ChiP-seq)
Chromatin Immunoprecipitation (CHIP) waa farsamo ka faa'iidaysata unugyada difaaca si ay si xushmad leh u xoojiso borotiinnada DNA-da ku xidha iyo bartilmaameedyada genomics ee u dhigma. Ku biirinteeda NGS waxay awood u siinaysaa sifaynta genome-ballaaran ee bartilmaameedyada DNA-da ee la xidhiidha wax ka beddelka histone, qodobbada qoraalka, iyo borotiinnada kale ee DNA-ku xidha. Habkan firfircoon ayaa awood u siinaya isbarbardhigga goobaha xidhitaanka ee noocyada unugyada kala duwan, unugyada, ama xaaladaha. Codsiyada ChIP-Seq waxay ka soo bilowdaan barashada hab-dhaqanka qoraalka iyo dariiqooyinka korriinka ilaa ay soo bandhigaan hababka cudurka, taasoo ka dhigaysa aalad lama huraan u ah fahamka hab-dhaqanka hab-nololeedka iyo horumarinta aragtiyada daaweynta.
Platform: Illumina NovaSeq
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Isku xigxiga genome bisulfite (WGBS)
Dhammaan Genome Bisulfite Sequencing (WGBS) waxay u taagan tahay habka heerka-dahabka ah ee sahaminta qoto dheer ee methyl DNA-da, gaar ahaan booska shanaad ee cytosine (5-mC), xakameynta muhiimka ah ee muujinta hiddaha iyo dhaqdhaqaaqa gacanta. Mabda'a ka hooseeya WGBS waxay ku lug leedahay daawaynta bisulfit, taasoo keenaysa beddelka cytosines-ka aan methylated ilaa uracil (C ilaa U), iyadoo ka tagaysa cytosines methylated oo aan isbeddelin. Farsamadani waxay bixisaa xallin hal-saldhig ah, taasoo u oggolaanaysa cilmi-baarayaasha inay si buuxda u baaraan methylome-ka oo ay ogaadaan qaababka methylation-ka ee aan caadiga ahayn ee la xiriira xaalado kala duwan, gaar ahaan kansarka. Shaqaalaysiinta WGBS, saynisyahannadu waxay heli karaan aragtiyo aan la qiyaasi karin oo ku saabsan muuqaallada methylation-ka ee genom-ballaaran, iyagoo siinaya faham qotodheer oo ku saabsan hababka epigenetic ee hoos yimaada hababka noole ee kala duwan iyo cudurrada.
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Qiimaynta chromatin-ka-wareejin-ka-hortagga leh ee la heli karo
ATAC-seq waa farsamo isku xigxig heersare ah oo loo isticmaalo falanqaynta gelitaanka chromatin-ballaaran ee genome-ga. Waxay adeegsataa waxay bixisaa faham qoto dheer oo ku saabsan hababka qalafsan ee xakamaynta epigenetic ee caalamiga ah ee muujinta hidda-wadaha. Habka wuxuu isticmaalaa transposase Tn5 firfircoon si uu isku mar u kala googooyo oo u sumadeeyo gobollada chromatin ee furan iyadoo la gelinayo iskuxirayaasha isku xigxiga. Kordhinta PCR ee soo socota waxay keenaysaa abuurista maktabad isku xigta, taas oo u oggolaanaysa aqoonsiga dhammaystiran ee gobollada chromatin-ka furan ee hoos yimaada xaalado waqti-meel gaar ah. ATAC-seq waxay bixisaa aragti guud ee muuqaalada chromatin ee la heli karo, si ka duwan hababka kaliya ee diirada saaraya goobaha xidhitaanka qodobka qoraalka ama gobolo gaar ah oo wax laga beddelay. Iyadoo la isku xigxigayo gobolladan chromatin-ka ah ee furan, ATAC-seq waxay muujinaysaa gobollada ay u badan tahay inay noqdaan taxane nidaamsan oo firfircoon iyo goobaha suurtagalka ah ee qoraalka qoraalka, iyagoo siinaya fikrado qiimo leh oo ku saabsan habaynta firfircoon ee muujinta hidda-socodka ee guud ahaan genome-ka.
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Taxanaha Bisulfite ee la dhimay (RRBS)
Matalaad la dhimay ee Bisulfite Sequencing (RRBS) ayaa u soo baxday sidii kharash-ool iyo wax-ku-ool ah beddelka Dhammaan Genome Bisulfite Sequencing (WGBS) ee cilmi-baarista methylation DNA. Iyadoo WGBS ay bixiso aragtiyo dhammaystiran iyadoo lagu eegayo genome-ga oo dhan hal xal oo sal ah, qiimaheeda sare wuxuu noqon karaa arrin xaddidaysa. RRBS waxay si xeeladaysan u yaraynaysaa caqabadan iyadoo si xushmad leh u falanqaynaysa qayb ka mid ah genome-ka. Habkani wuxuu ku tiirsan yahay kobcinta gobollada jasiiradaha hodanka ah ee CpG iyadoo la kala gooyo MspI oo ay ku xigto cabbirka xulashada 200-500/600 bps jajab. Sidaa darteed, gobollada u dhow jasiiradaha CpG oo keliya ayaa la isku xig-xigay, halka kuwa jasiiradaha CpG fog leh laga saaray falanqaynta. Nidaamkan, oo ay weheliso isku-dhafka bisulfit, wuxuu u oggolaanayaa in la ogaado xallinta sare ee methylation DNA, iyo habka isku xigxiga, PE150, ayaa si gaar ah diiradda u saaraya cidhifyada gelinta halkii dhexe, kordhinta waxtarka methylation profile. RRBS waa qalab aad u qiimo badan oo awood u siinaya cilmi-baarista methylation DNA ee qiimaha-wax-ku-ool ah iyo horumarinta aqoonta hababka epigenetic.
